Dissolution mechanism of supported phospholipid bilayer in the presence of amphiphilic drug investigated by neutron reflectometry and quartz crystal microbalance with dissipation monitoring.

The influence and interaction of the ionizable amphiphilic drug amitriptyline hydrochloride (AMT) on a 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) phospholipid bilayer supported on a silica surface have been investigated using a combination of neutron reflectometry and quartz crystal microbalance with dissipation monitoring. Adding AMT solutions with concentrations 3, 12, and 50 mM leaves the lipid bilayer mainly intact and we observe most of the AMT molecules attached to the head-group region of the outer bilayer leaflet. Virtually no AMT penetrates into the hydrophilic head-group region of the inner leaflet close to the silica surface. By adding 200 mM AMT solution, the lipid bilayer dissolved entirely, indicating a threshold concentration for the solubilization of the bilayer by AMT. The observed threshold concentration is consistent with the observation that various bilayer structures abruptly transform into mixed AMT-DOPC micelles beyond a certain AMT-DOPC composition. Based on our experimental observations, we suggest that the penetration of drug into the phospholipid bilayer, and subsequent solubilization of the membrane, follows a two-step mechanism with the outer leaflet being removed prior to the inner leaflet.

Orientation Distribution of Molecules in a Smectic Liquid Crystal with a Distorted Director Geometry.

Quantitative determination of the molecular orientation distribution function in samples of liquid crystals with a complex director geometry was performed using the numerical simulation of electron paramagnetic resonance (EPR) spectra of the spin probes in a liquid-crystalline medium. To achieve the quantitative agreement of experimental and simulated EPR spectra, the hierarchy of the orientation order was explicitly taken into account, namely, the local ordering of liquid crystal molecules by the mean-field potential of surrounding molecules, and the partial disordering of local directors within the sample. The samples under study are planar and twist LC cells with liquid crystal 8CB cooled from the nematic into the smectic A phase in the magnetic field. The presence of the magnetic field perpendicular to the cell director leads to distortion of the orientation of the liquid crystal in the cell. The spin probe technique was successfully employed for the reliable measurement of orientation distribution functions of the low nonorthorhombic symmetry. Orientation order parameters up to 12th rank were measured, including nonaxial and nonorthorhombic order parameters. It is shown that the presence of several contradicting aligning forces leads to the tilt of the preferential director toward the direction, which is a compromise between the orienting forces.

A Digital Electrocardiographic System for Assessing Myocardial Electrical Instability: Principles and Applications.

The aim of the study was to develop an ECG hardware and software system for monitoring electrical instability of the myocardium and to assess the diagnostic and prognostic capabilities of this setup in a cardiology clinic. MATERIALS AND METHODS: The Intecard 7.3 software and hardware system developed in this study makes it possible to measure fluctuations of the ECG amplitude-time parameters using the beat-to-beat mode. Intecard 7.3 evaluates a number of ECG markers that reflect electrical instability of the myocardium. Among them are the fragmented QRS complex, the spatial QRS-T angle, the T-wave alternans, the duration, and dispersion of the QT interval, the turbulence and acceleration/deceleration of the heart rhythm.Clinical trials of Intecard 7.3 were carried out with 734 patients with ischemic heart disease or cardiomyopathy and 112 healthy individuals. RESULTS: Intecard 7.3 reliably identifies fragmented QRS complexes by detecting short spikes of <25 ms in the ascending parts of the Q, R, and S waves. The QRS-T angle is determined from the reference amplitudes of the R and T waves in leads avF, V2, V5, and V6. Digital precision processing of the ECG signal improves its accuracy to microvolts and microseconds.The software was designed to measure the T-wave amplitude in each of 300-500 cardiobeats; T-wave alternans was estimated by the moving average method. In a typical cardiobeat, the QT dispersion was calculated based on 12 ECG leads. From the sequence of RR intervals, turbulence, and deceleration of the heart rhythm were determined.During the observation period of 5.0 [2.1; 5.9] years, 90 out of 734 patients (12.3%) experienced adverse cardiovascular events (ACVE). In this period, the myocardial electrical instability was recorded in patients with ACVE more frequently than in those without ACVE. Thus, the frequency of fragmented QRS was 72.2±4.7 vs 16.8±1.5% (p<0.01), the values of the QRS-T angle were 128 [55; 101] vs 80 [53; 121]° (p<0.001), the T-wave alternans - 36.9 [15.5; 62.1] vs 21.9 [10.2; 30.7] µV (p<0.005), the QT interval - 408 [383; 438] vs 376 [351; 400] ms (p<0.001), the QT dispersion - 76 [57; 96] vs 64 [50; 92] ms (p<0.005), respectively. In patients with ACVE, the threshold that triggers pathological rhythm turbulence was higher (>0%) than that in healthy controls (p<0.001); the deceleration of the heart rhythm was reduced from 19.2 [2.2; 38.0] to 8.8 [4.0; 16.8] ms (p<0.05).A personalized model for ACVE risk stratification has been developed. In this model, the area under the ROC curve was 0.856; sensitivity - 75%; specificity - 78%; predictive accuracy - 77%. CONCLUSION: Using the ECG markers of myocardial electrical instability, the Intecard 7.3 system allows one to predict life-threatening ventricular tachyarrhythmias and sudden cardiac death with an accuracy of 77%. The non-invasiveness, high productivity, and reasonable cost ensure the availability of this predictive technology in all levels of healthcare.

Effect of Surgical Technique on the Microstructure and Microcirculation of the Small Intestine Stump during Delayed Anastomosis: Multimodal OCT Data.

The aim of the study was to use multimodal optical coherence tomography (MM OCT) to evaluate microstructure and microcirculation in the proximal and distal sections of the intestine relative to the resected area in acute mesenteric ischemia. MATERIALS AND METHODS: The study was carried out using three groups of male Wistar rats weighing 270-435 g (n=18). Acute occlusive arterial ischemia of the small intestine was initiated in all animals. After 80-90 min, the ischemic non-viable part of the intestine was resected, and the operation was completed by stoma exteriorization (group 1, n=6), by applying purse-string sutures (group 2, obstructive resection, n=6), or by internal drainage of the proximal and distal ends of the small intestine (group 3, bypass, n=6). Relaparotomy and anastomosis formation were performed 2 days later.With the help of MM OCT at each stage of the surgical intervention, images were obtained from the serous membrane side: the intestinal wall microstructure (layers) was viewed using cross-polarization OCT (CP OCT) and the intramural circulation - using optical coherent angiography (OCA). The MM OCT images obtained from the terminal intestine sections immediately after resection and 2 days later (before the anastomosis formation) were compared between the experimental groups, as well as with the pre-ischemic data (norm). All resected sections of the intestine were then histologically examined. The MM OCT data were compared with the histological and intravital macroscopy data. RESULTS: As a result of studying the intestinal wall microstructure by in vivo CP OCT, it was found that during ostomy (group 1) and obstructive resection (group 2), the images showed signs of tissue edema and destructive changes in the mucous membrane that were confirmed histologically, while with bypass surgery (group 3), there were minimal changes as compared with the norm.According to the OCA data, on day 2 of ostomy in the proximal and distal segments of the intestine, there was a noticeable disappearance of small and medium blood vessels; mainly large arteries and veins could be visualized. Following obstructive resection (purse-string suturing) or bypass surgery, the most noticeable changes (a decrease in the number of visualized blood vessels) were observed in the distal part of the intestine. The L index calculated from OCA images and characterizing the total length of the intramural perfused vasculature, showed a statistically significant decrease during ostomy: 12.18 [10.40; 14.20] µm - in the proximal and 10.67 [7.98; 13.05] µm - in the distal section; for comparison, the L index before ischemia was 18.90 [17.98; 19.73] µm and 18.74 [17.46; 19.90] µm, respectively (p=0.0001). In obstructive resection (group 2), statistically significant differences in the L parameter were found only for the distal bowel section: 16.39 [12.37; 18.10] µm compared with 18.74 [17.46; 19.90] µm before ischemia (p=0.041). After bypass surgery (group 3), there were no significant deviations in the L index. CONCLUSION: By using MM OCT, we found that in treating the remaining sections of the intestine after its emergency resection for acute mesenteric ischemia, the type of surgical technique determines the tissue structure in the period before the delayed anastomosis is applied.The least pronounced and most balanced changes occur in the proximal and distal segments of the intestine when operated using the bypass technique. However, to recommend this type of surgery, the development of reliable, safe, and effective bypass instruments is needed.

Rotational Dynamics of Nitroxide Biradical in Room-Temperature Ionic Liquids Measured by Quantitative Simulation of EPR Spectra.

Temperature dependences of electron paramagnetic resonance (EPR) spectra of an imidazoline nitroxide biradical spin probe in a series of room-temperature ionic liquids in the temperature range 124-390 K have been quantitatively simulated. The unusual asymmetric EPR spectrum shape previously observed in these systems [Kokorin et al., Appl. Magn. Res. 48 (2016) 287] is shown to originate from anisotropic rotational diffusion of the probe molecule. All experimental spectra were quantitatively reproduced in simulation using a unified set of geometrical and magnetic parameters of the spin probe, which were found to be fully consistent with the biradical geometry obtained from density functional theory calculations. Temperature dependences of rotation diffusion coefficient of the probe characterize the molecular mobility of the ionic liquid, whereas the temperature dependences of the spin-exchange integral J and of the isotropic hyperfine interaction constant, aN, are shown to reflect the intramolecular conformation motions of the biradical probe.

[Surgical treatment of epilepsy in children with focal cortical dysplasia]. / Khirurgicheskoe lechenie epilepsii u detei s fokal'noi kortikal'noi displaziei.

OBJECTIVE: Surgery is the first-line treatment option in children with FCD and refractory epilepsy, but the rate of success and patient numbers who became free of seizures vary widely from series to series. STUDY AIMS: To elicit variables affecting the outcome and predicting achievement of the long-term seizure-free status. MATERIAL AND METHODS: One hundred sixty-nine children with cortical dysplasia and DR-epilepsy underwent surgery Preoperative evaluation included prolonged video-EEG and MRI (in all patients) and neuropsychological testing when possible. Fourteen patients underwent invasive EEG, fMRI and MEG were used also in some cases. Including 27 repeat procedures the list of overall 196 surgeries performed consists of: cortectomy (lesionectomy with or without adjacent epileptogenic cortices) ­ in 116 cases; lobectomy ­ in 46; and various disconnective procedures ­ in 34 patients. Almost routinely employed intraoperative ECOG (134 surgeries) was combined with stimulation and/or SSEP in 47 cases to map eloquent cortex (with CST-tracking in some). A new permanent and not anticipated neurological deficit developed post-surgery in 5 cases (2,5%). Patients were follow-upped using video-EEG and MRI and FU which lasts more than 2 years (median ­ 3 years) is known in 56 cases. Thirty-two children were free of seizures at the last check (57,2% rate of Engel IA). A list of variables regarding patients' demography, seizure type, lesion pathology and localization, and those related to surgery and its extent were evaluated to figure out anyone associated with favorable outcome. RESULTS: Both Type II FCDs and their anatomically complete excision are positive predictors for favorable outcome and achievement of SF-status (p<0,05). Residual epileptic activity on immediate post-resection ECOG do not affect the outcome. CONCLUSION: Patients with Type II FCD, particularly with Type IIb malformations are the best candidates for curative surgery, including cases with lesions in brain eloquent areas. Kids with Type I FCD have much less chances to become free of seizures when attempting focal cortectomy. However, some of them with early onset catastrophic epilepsies may benefit from larger surgeries using lobectomy or various disconnections.

[Pathogenesis, diagnosis, prevention and treatment of disseminated intravascular coagulation syndrome in COVID-19 infection].

AIM: Clinical characteristics of disseminated intravascular coagulation (DIC) in COVID-19 infection and assessment of the effectiveness of complex therapy for this syndrome at the stages of prevention and treatment of various complications. MATERIALS AND METHODS: The study of publications was carried out through search engines on the Internet using keywords. To diagnose the infection, the COVID-19 program was used on the MeDiCase platform, which is publicly available on www.medicase.pro, which suggests a diagnosis with a sensitivity of 89.47%. The study included 85 patients with acute COVID-19 with mild to moderate disease, aged 11 to 81 years. The presence of the pathogen was confirmed immunologically in 12% of patients; in other cases, the diagnosis was based on the results of an automated survey in the MeDiCase system. All patients, according to the MGNOT recommendations, were prescribed one of the oral direct anticoagulants - Eliquis at a dose of 5 mg 2 times a day, Ksarelto at a dose of 10 mg 2 times a day or Pradax at a dose of 110 mg 2 times a day for at least 2 weeks. All other drugs with antiviral, immunomodulatory effects, antibiotics were canceled. RESULTS: The presence of DIC is substantiated by the morphological picture of changes in organs and tissues, clinical (hematoma-petechial type of bleeding in combination with thromboembolic syndrome and the presence of thrombovasculitis) and laboratory changes: an increase in the level of soluble fibrin-monomer complexes, D-dimer, hyperfibrinogenaemia, less often - thrombocytopenia, violation of fibrinolytic activity. The phenomenon of consumption of clotting factors and profuse bleeding are rare. Direct anticoagulants, fresh frozen plasma transfusions and plasmapheresis are used in the treatment of disseminated intravascular coagulation. The paper presents its own positive results of early prescription at the outpatient stage of direct oral anticoagulants in prophylactic doses (no case of disease progression), individual cases of the use of fresh frozen plasma and plasapheresis. CONCLUSION: DIC syndrome with the development of thrombovasculitis is the most important pathogenetic mechanism for the development of microthrombotic and hemorrhagic disorders in organs during infection with COVID-19, leading to dysfunction of the lungs, brain and other nerve tissues, kidneys, thromboembolic complications, etc. Many symptoms of the disease may be associated with a violation of the nervous regulation of the functions of organs and systems. Prevention of thrombovasculitis is effective already at the stage of the first manifestation of the disease with the outpatient use of direct anticoagulants (oral, low molecular weight heparins). In case of more severe manifestations (complications) of the disease, additional use of freshly frozen plasma and plasmapheresis is effective.

Spin Probe Determination of Molecular Orientation Distribution and Rotational Mobility in Liquid Crystals: Model-Free Approach.

A model-free approach for simulation of EPR spectra of nitroxide spin probes in liquid-crystalline materials was suggested and used to obtain parameters of molecular orientation and rotational mobility. The developed method is based on experimental recording and numerical simulation of the angular dependence of EPR spectra, which is shown to be much more informative in comparison with a single EPR spectrum. Quantitative spectral simulations considering both local orientational ordering and distribution of local directors in the sample were used for discrimination of models of rotational mobility and orientational alignment. The method was applied for detailed quantitative characterization of axial, orthorhombic, and low-symmetry non-orthorhombic molecular orientation distributions. It is shown that the ordinarily used model of rotational diffusion in a mean-field potential is suitable for the description of molecular mobility and orientational ordering only for relatively low sample temperatures and low-mobility probe molecules with large sizes. In cases of high molecular mobility, the more realistic jump mechanism of molecular moves can be approximately described as quasi-librations. For ordered liquid crystals it was found that mostly the order parameters up to rank 12-14 are essential and easily determined. When well-aligned materials are described, the order parameters up to 18th rank or even higher become meaningful. Both molecular and sample biaxiality is analyzed and quantitatively characterized. The local molecular ordering and sample orientational alignment are quantitatively characterized separately.

[Laparoscopic ultrasound guided partial nephrectomy].

INTRODUCTION: The choice of method of surgical treatment for endophytic kidney tumors depends on the precise definition of the location and size, the extent of invasion into adjacent tissues and the type of vascularization. Intraoperative ultrasound (IOUS) in combination with 3D-CT allow to receive this information. The aim of this study was to compare the laparoscopic-assisted extracorporeal partial nephrectomy and laparoscopic partial nephrectomy with IOUS and visualization in Full HD, 3D Full HD and 4K modes. MATERIALS AND METHODS: A total of 77 patients aged 43-75 years with endophytic renal tumors were included in the study. They were undergone either extracorporeal partial nephrectomy or laparoscopic partial nephrectomy with IOUS. We compared the rate of positive surgical margins, early postoperative bleeding, de novo renal failure or aggravation of preexisting renal failure and stricture of vesico-ureteric anastomosis. RESULTS: The rate of early postoperative bleeding, chronic kidney failure and pathologically-proven positive surgical margin in patients who underwent extracorporeal partial nephrectomy followed by kidney autotransplantation was 23.1%, 13.4% and 13.4%, respectively. After laparoscopic partial nephrectomy these values were 12%, 16.7% and 8.3%, respectively. The best outcomes were achieved in patients who underwent laparoscopic partial nephrectomy with IOUS and with a use of visualization in Full HD, 3D Full HD and 4K modes.

[The role of translocations involving c-MYC/8q24, BCL2/18q21 and/or BCL6/3q27 genes in patients with follicular lymphoma. Retrospective analysis of single - centre data].

Aim of the issue was to compare clinical characteristics and treatment results of patients with follicular lymphoma (FL) with translocations involving loci of c-MYC/8q24, BCL2/18q21 and/or BCL6/3q27 genes and patients with high - grade B-cell lymphoma [High - grade B-cell lymphoma (HGBL), double - hit (DH)]. Materials and methods. Since 2004 to 2017 years in National Research Center for Hematology 12 patients with high - grade B-cell lymphoma double - hit (HGBL DH) and 6 FL patients with translocations involving c-MYC and BCL2 and/or BCL6 had been treated. We performed a comparative analysis of clinical characterisctics in both groups. As primary endpoints was assessed frequency of complete remission (CR) or progressive disease (PD); as secondary endpoints - overall (OS) and event - free survival (EFS). Results. 5 patients with HGBL DH had c-MYC/BCL6, 7 - c-MYC/BCL2 rearrangements; 2 patients with FL had c-MYC/BCL2, 3 - c-MYC/BCL6, 1 - c-MYC/BCL2/BCL6 rearrangements. FL was represented by grade 3A in 2, grade 3B - in 4 cases, 3 of them had large - cell transformation. In HGBL DH and FL patients had no significant differences in clinical characteristics. The majority of patients had a widespread tumour, increased LDH activity, high frequency of extranodal and bone marrow involvement. Ki-67 expression level was lower in patients with FL (p.

[Left-sided renal cell cacrinoma with vena cava inferior thrombus: totally laparoscopic removal].

A surgical treatment of patients with tumor thrombus in the inferior vena cava (IVC) of levels II-III, originating from the left renal vein involves performing thrombectomy, radical nephrectomy and lymph nodes dissection. In most cases it requires major open surgery which leads to complications in 38% of patients and perioperative mortality of 4-10%. In recent years, the laparoscopic radical nephrectomy with thrombectomy have been gradually introduced. However, there are anecdotal reports about performing of such interventions in case of left-sided renal tumor with thrombus in the IVC. A description of technique and own experience of performing laparoscopic radical nephrectomy with thrombectomy from the IVC in 3 patients with tumor thrombus of levels II-III, originating from left kidney are presented. In one case distant metastases were detected preoperatively and in another patient an involvement of tail of the pancreas by the tumor was diagnosed. There was no conversion to open surgery. The maximum tumor size ranged from 5 to 16 cm. The length of tumor thrombus in the IVC was 2.4-7 cm and estimated blood loss was 300-2500 ml. In one case a blood transfusion was required postoperatively. The follow-up period was 4-26 months. One patient died from progression of the disease after 5 months, two other patients are alive without any signs of relapse. The initial experience of laparoscopic radical nephrectomy with thrombectomy from IVC in patients with left-sided renal cell cancer suggests that this technique is reproducible and relatively safe with respecting of basic principles of oncology and vascular surgery. The surgery is not associated with major perioperative complications, significant blood loss and is accompanied by a rather favorable postoperative period. An accumulation of a larger number of cases is needed to assess the oncological results of this technique.

Approaches to Controlled Co-Amplification of Genes for Production of Biopharmaceuticals: Study of the Insertion and Amplification Dynamics of Genetic Cassettes in the Genome of Chinese Hamster Ovary Cells during Co-Expression of Compatible Pair of Plasmids.

Plasmid vector family p1.1 based on non-coding regions of Chinese hamster housekeeping gene EEF1A and concatemer of Epstein-Barr virus terminal repeat increases the frequency of genome integration and provides rapid amplification of the target genes in the genome. For a pair of fluorescent proteins eGFP and mCherry it was shown that p1.1 vectors bearing dihydrofolate reductase and glutamine synthetase selection markers upon co-transfection into CHO DG44 cell line allow obtaining a polyclonal cell population in which ~70% of cells express both genes. The subsequent one-step gene amplification of the genome-integrated genetic cassettes under the selective pressure of increased concentrations of methotrexate can increase the expression of both integrated genes up to 8.2% eGFP and 9.9% mCherry of total protein. This approach can be used for the development of cell lines for the production of functional heterodimeric proteins, e.g. polypeptide hormones and therapeutic antibodies.

Substantial Difference in Ordering of 10, 15, and 20 nm Iron Oxide Nanoparticles on a Water Surface: In Situ Characterization by the Grazing Incidence X-ray Scattering.

In the present study, for the first time, a unique combination of in situ grazing incidence small-angle X-ray scattering and X-ray reflectivity, accompanied by the pressure-area isotherm analysis, Brewster angle microscopy, and ex situ scanning electron microscopy, was applied for investigation of two-dimensional superlattices of magnetic nanoparticles as they form on a water surface in a Langmuir trough. Iron oxide particles of different sizes stabilized with a single layer of oleic acid were used. It is demonstrated that monodisperse 10 nm particles on a water surface reproducibly form identical highly ordered monolayers in a wide range of experimental conditions, while monodisperse 20 nm particles always form compact three-dimensional clusters and never the monolayers. Monodisperse particles of an intermediate size, 15 nm in diameter, build a metastable monolayer, which shows a tendency for spontaneous transformation to bi-, tri-, and multilayer islands. The importance to use both grazing incidence small-angle X-ray scattering and X-ray reflectivity together with the complementary techniques, to avoid misinterpretation of separate experimental data sets, is underlined.

[CYTOREDUCTIVE NEPHRECTOMY IN METASTATIC RENAL CELL CANCER].

The role of cytoreductive nephrectomy in the current era of targeted therapy remains unknown. Two prospective randomized phase III trials (CARMENA and SURTIME) are now opened to evaluate the efficacy of cytoreductive nephrectomy in patients with metastatic renal cell carcinoma. So far it is not well known who will and who will not benefit from such surgery.

Molecular orientational order of nitroxide radicals in liquid crystalline media.

The orientational distribution of a set of stable nitroxide radicals in aligned liquid crystals 5CB (nematic) and 8CB (smectic A) was studied in detail by numerical simulation of EPR spectra. The order parameters up to the 10th rank were measured. The directions of the principal orientation axes of the radicals were determined. It was shown that the ordering of the probe molecules is controlled by their interaction with the matrix molecules more than the inherent geometry of the probes themselves. The rigid fused phenanthrene-based (A5) and 2-azaphenalene (A4) nitroxides as well as the rigid core elongated C11 and 5α-cholestane (CLS) nitroxides were found to be most sensitive to the orientation of the liquid crystal matrixes.

Determination of structural characteristics of all-organic radical liquid crystals based on analysis of the dipole-dipole broadened EPR spectra.

The angular dependences of g-value and line width of EPR spectra of paramagnetic all-organic liquid crystalline (LC) materials were measured for the quantitative characterization of the nematic, cholesteric, and smectic C phases. The detailed molecular alignment in mesophases was determined by means of numerical spectra simulation focusing on spin exchange and dipole-dipole magnetic interactions of neighboring molecules. The obtained structural data indicate that the spin polarization mechanism between neighboring molecules rather than the direct through-space interactions between paramagnetic centers is responsible for the specific magnetic properties of the studied LC materials.

Blood Clotting Factor VIII: From Evolution to Therapy.

Recombinant blood clotting factor VIII is one of the most complex proteins for industrial manufacturing due to the low efficiency of its gene transcription, massive intracellular loss of its proprotein during post-translational processing, and the instability of the secreted protein. Improvement in hemophilia A therapy requires a steady increase in the production of factor VIII drugs despite tightening standards of product quality and viral safety. More efficient systems for heterologous expression of factor VIII can be created on the basis of the discovered properties of its gene transcription, post-translational processing, and behavior in the bloodstream. The present review describes the deletion variants of factor VIII protein with increased secretion efficiency and the prospects for the pharmaceutical development of longer acting variants and derivatives of factor VIII.

Stable Expression of Recombinant Factor VIII in CHO Cells Using Methotrexate-Driven Transgene Amplification.

Prophylaxis and treatment of inherited clotting disorder hemophilia A requires regular administration of factor VIII. Recombinant factor VIII, which is produced in CHO or BHK cells, is equivalent to the plasma-derived one and is prevalent in current clinical practice in developed countries. Development of a biosimilar recombinant FVIII requires the creation of a highly productive clonal cell line and generation of monoclonal antibodies suitable for affinity purification of the product. Methotrexate-driven transgene amplification of genetic cassettes that code full-length and truncated variants of FVIII under the control of the CMV promoter was studied. It was shown that the expression level of the truncated variant of FVIII is 6.5 times higher than that of the full-length molecule. The transgene amplification procedure was sufficient for a twofold increase of the expression level in the transfected cells pool and subsequent selection of the clonal line, stably producing truncated FVIII at the level of 0.52 IU/ml during cultivation in a chemically defined protein-free culture medium. Four generated mouse monoclonal antibodies toward the heavy chain of FVIII were found suitable for binding the truncated variant of FVIII directly from the conditioned medium and elution of the FVIII with a more than 85% yield and normal pro-coagulant activity. The producer cell line and monoclonal antibodies obtained are sufficient for the development of upstream and downstream processes of biosimilar FVIII production. Generation of more productive cell lines by the use of stronger, nonviral promoters and shorter cDNA of FVIII will be the subject of further studies.

Order parameters , , and of aligned nematic liquid-crystalline polymer as determined by numerical simulation of electron paramagnetic resonance spectra.

High rank order parameters may comprise substantial information about molecular orientational distribution of liquid-crystalline materials. There are few experimental procedures targeted at the determination of high rank order parameters. We suggest a procedure for the determination of order parameters of a spin probe by numerical simulation of electron paramagnetic resonance (EPR) spectra. The procedure is based on revealing the molecular orientation axis of a spin probe. Order parameters , , and of the spin probe in aligned liquid-crystalline polymer have been measured by the suggested procedure. It was shown that the procedure gives more reliable and complete characteristics of molecular order than does polarized UV-visible spectroscopy.

Self-assembled iron oxide nanoparticle multilayer: x-ray and polarized neutron reflectivity.

We have investigated the structure and magnetism of self-assembled, 20 nm diameter iron oxide nanoparticles covered by an oleic acid shell for scrutinizing their structural and magnetic correlations. The nanoparticles were spin-coated on an Si substrate as a single monolayer and as a stack of 5 ML forming a multilayer. X-ray scattering (reflectivity and grazing incidence small-angle scattering) confirms high in-plane hexagonal correlation and a good layering property of the nanoparticles. Using polarized neutron reflectivity we have also determined the long range magnetic correlations parallel and perpendicular to the layers in addition to the structural ones. In a field of 5 kOe we determine a magnetization value of about 80% of the saturation value. At remanence the global magnetization is close to zero. However, polarized neutron reflectivity reveals the existence of regions in which magnetic moments of nanoparticles are well aligned, while losing order over longer distances. These findings confirm that in the nanoparticle assembly the magnetic dipole-dipole interaction is rather strong, dominating the collective magnetic properties at room temperature.